C. parvum is an enteric protozoan that infects the gastrointestinal tract of most mammalian species and is the major cause of cryptosporidiosis in humans. C. Parvum comprises two known genotypes, type 1 and type 2. Type 1 (anthroponotic) is found exclusively in humans, while type 2 (zoonotic) is found in all mammals including humans. C. meleagridis (which infects birds and mammals) has also been associated with human cryptosporidiosis recently. C. Parvum type 1 is responsible for the majority of human infections and differs from type 2 in many aspects. Evidence from animal studies in this laboratory and in Ugandan children indicates that types 1 and 2 display several reproducibly distinct phenotypic traits including sporozoite proteins. Type 2 infects rodents but not type 1. Co-infections of animals with both types show no evidence of genetic recombination, and, therefore, it is likely that they maintain independent reproductive cycles. In contrast, recombination among type 2 isolates has been demonstrated experimentally. In addition, significant differences in DNA and amino acid sequences of types 1 and 2 surface protein gp40/IS exist. These observations suggest that the two types exhibit prominent genotypic and phenotypic differences and consequently, may belong to separate species of Cryptosporidium. While C. parvum types 1 and 2 and C. meleagridis display differences in host range, they share a common denominator in that all three appear to infect humans. Several type I isolates have recently been successfully adapted to continuously propagate in gnotobiotic piglets, enabling for the first time to begin studies on type 1 isolates and compare them with what is already know about type 2 isolates. In this application, the investigators aim to exploit the piglet model to define differences, if any, between C. parvum types 1 and 2. C. meleagridis (the only known species to cross the vertebrate class barrier) has also been successfully adapted to propagate continuously in piglets and will be included in this comparative study, largely because of its major epidemiologic and public health significance. The long-term goal of the candidate's research is to characterize molecular, functional, and immunologic parasite antigens associated with virulence and protection. The proposed study is the first step in that direction and the outcomes will provide basis for the future studies. The following are several hypotheses that will be tested in this study: 1) as observed in human volunteer studies, there will be differences in virulence among isolates of type 1, 2, and C. meleagridis, in terms of clinical manifestation, infectivity, and the extent and distribution of the mucosal damage (Aim 1); and 2) while the nature of systemic and mucosal immune responses are likely to be almost identical among the three types/species of Cryptosporidium, immunodominant antigenic differences will be identified (Aims 2 and 4) which will result in greater cross-protection within homologous isolates than heterologous, and that C. meleagridis will be considerably different than either type (Aim 3). The following specific aims are designed to address these hypotheses: 1) compare the nature of host parasite interaction of C. parvum types 1 and 2, and C. meleagridis in the gnotobiotic piglet model with respect to disease manifestation and pathogenesis; 2) characterize systemic as well as local cellular and humoral immune responses against C. parvum types 1 and 2 and C. meleagridis in the gnotobiotic piglet model; 3) determine the extent of cross-protection between C. parvum types 1 and 2 and C. meleagridis in the gnotobiotic piglet model; and 4) identify immunodominant molecules that are unique to each C. parvum type and C. meleagridis, and also that are common among C. parvum types and C. meleagridis, which will help in identification of key specific- and cross-protective parasite antigens.